In humans, chronically increased signaling through Gi-coupled receptors is associated with congestive heart failure (CHF) caused by idiopathic dilated cardiomyopathy or ischemic cardiomyopathy following myocardial infarction. However, the mechanisms responsible are unclear. Our working hypothesis is that chronically increased Gi signaling causes impaired excitation-contraction (ec) coupling. To test this hypothesis we will combine physiological measurements of cardiac muscle function with a novel transgenic mouse model in which a modified Gi-coupled receptor (Ro1) is targeted to the heart. Expression of Ro1 is regulated by a tetracycline-controlled expression system (tet-system). We have recently shown that chronic Ro1 expression causes CHF and major abnormalities of Ca2+ transients and contraction. In contrast, acute Ro1 expression causes significant protection against ischemia/reperfusion injury, suggesting a dual role for increased Gi signaling in cardioprotection and disease. For this proposal we will determine the ec-coupling mechanisms and Gi signaling mechanisms involved in CHF and cardioprotection. Using single myocytes, cardiac trabeculae, and Langendorff perfused mouse hearts, we will determine the effect of Ro1 expression on Ca2+ transients and determine the mechanisms responsible by localizing abnormalities to specific Ca2+ handling processes. We will determine the effect of Ro1 expression on Ca2+-responsiveness and determine the mechanisms responsible by localizing abnormalities to specific contractile and regulatory proteins. Using the tet-system to turn off Ro1 expression after induction of CHF, we will determine the extent to which ec-coupling abnormalities are reversible. To elucidate signaling mechanisms, we will determine which of the major Gi pathways in the heart (Gi2 and Gi3) are involved; and whether signaling via the G protein alpha subunit and/or the betagamma dimer is involved. Using 3 model systems we will investigate Gi signaling effects (both deleterious and beneficial) and the ec-coupling- and signaling mechanisms involved in: Aim 1. CHF caused by Ro1 expression; and recovery after terminating Ro1 expression. Aim 2. Acute Cardioprotection caused by Ro1 expression. Aim 3. CHF caused by ischemic cardiomyopathy. This research will provide new information on the dual role of Gi signaling in both heart failure and cardioprotection which may help identify new strategies to treat heart disease.